The purpose of this project is to characterize second messenger pathways involved in receptor regulation of neurotransmitter levels. Previous work from our lab demonstrated that angiotensin II (AII) and nicotine each stimulate enkephalin secretion and gene expression in cultured bovine chromaffin cells. Both agonists increase intracellular CA2+ concentration (Cai) and protein kinase C (PKC) translocation through different mechanisms of action. Nicotine activates cation channels, 1-type Ca2+ channels in particular being important for both Cai and PKC responses; the latter response is long-lived and may contribute to enkephalin gene regulation. AII activates phospholipase C (PLC) leading to mobilization of intracellular CA2+; AII also activates CA2+ influx partly through omega- Cgtx-sensitive channel which is critical for secretory effects. AII is ineffective as a secretagogue after inhibition or activation of K+ channels, suggesting that effects are mediated on Cgtx- and (voltage-) sensitive Ca2+ channels via inhibiting K+ channels. We hope that these agonists will be models for excitatory amino acid agonists in rat brain cell cultures. Cultured striatal cells show PKC responses to cation channel-linked receptors-- NMDA, kainate and AMPA-- and to PLC-linked receptor agonists-- taCPD and dopamine and norepinephrine. Dopamine and AMPA responses are dependent on the age of the rat used. The contribution of these agonists, and the CA2+ and PKC pathways to enkephalin expression in brain cultures is presently being assessed.